Increased oxidative stress in dopaminergic neurons has been implicated as a causative factor in Parkinson's disease (PD). The catechols DOPA dopamine and DOPAC generated in these cells can undergo oxidation to produce aggressive oxygen species and reactive arylating quinones which are capable of cellular damage. We have characterized a null polymorphism in the NQO1 gene (NQO1 *2) and individuals carrying the variant NQO1 *2 allele have been found to be at a markedly increased risk of PD relative to control individuals emphasizing the role of NQO1 as a risk factor for PD. PD has also been associated with inhibition of the ubiquitin-proteasomal system (UPS). Inhibition of the UPS by aggregated proteins in neurodegenerative disease leads to increased oxidative stress. Since oxidative stress can result in the misfolding and aggregation of proteins resulting in further frustration of the UPS, a picture is beginning to emerge where both oxidative stress and UPS inhibition are part of a vicious cycle and are important factors in the etiology of PD.In this proposal, we will test the following hypotheses; 1) that NQO1 protects cells against reactive arylating quinones and oxidizing species generated during metabolism of DOPA, dopamine and DOPAC. We will use dopaminergic cells, mechanism based inhibitors of NQO1 and an isogenic cellular model developed in our lab to explore the role of NQOl in these studies; 2) that DOPA, dopamine or DOPAC derived o-quinones contribute to UPS inhibition and that NQO1 protects against this inhibitory effect; 3) that NQO1 protects against oxidative stress generated as a result of inhibition of the UPS. UPS inhibition in cells will be achieved by the use of both chemical proteasome inhibitors and by transfection of mutant forms of alpha-synuclein which have been associated with PD and inhibit the UPS by mechanisms involving protein aggregation and 4) that mutant NQO1*2 protein, which is normally rapidly degraded by the UPS, generates oxygen radicals leading to increased oxidative stress. The ability of mutant NQO1 *2 protein to generate oxygen radicals becomes particularly important under conditions where the UPS is inhibited and the protein accumulates.